本文選題：玉米 + 花藥角質(zhì)層��； 參考：《中國農(nóng)業(yè)大學(xué)》2017年博士論文

【摘要】：玉米(Zea mays)是一種重要的糧食作物和經(jīng)濟(jì)作物。雄性不育系在玉米雜交種子生產(chǎn)中具有重要應(yīng)用價值�；ㄋ幗琴|(zhì)層和花粉外壁是花粉形成的兩類保護(hù)性結(jié)構(gòu)。在擬南芥(Arabidopsis thaliana)和水稻(Oryza sativa)中,部分控制花藥角質(zhì)層和花粉外壁發(fā)育的基因已被克隆。然而,在玉米中僅有少數(shù)幾個基因被克隆,且它們的分子機(jī)制尚不清楚。通過篩選實(shí)驗(yàn)室Mutator突變體庫,我們獲得了一份雄性不育系-ipe1,應(yīng)用圖位克隆和分子生物學(xué)等方法,對IPE1基因進(jìn)行了克隆并初步闡明了它調(diào)控玉米育性的分子機(jī)制,主要研究結(jié)果如下:(1)表型分析表明ipe1僅花藥發(fā)育異常,其余器官發(fā)育無顯著變化;與野生型相比,散粉期ipe1花藥不能從穎殼露出,變小,萎蔫;碘-碘化鉀染色證明ipe1花藥無成熟花粉粒形成。(2)細(xì)胞學(xué)分析發(fā)現(xiàn)ipe1花藥角質(zhì)層和烏氏體形成受阻;突變體減數(shù)分裂是正常的,而柱狀層發(fā)育異常導(dǎo)致孢粉素隨機(jī)堆積和花粉粒外壁不規(guī)則,從而使小孢子從單核中期開始逐步降解;絨氈層相對于野生型降解提前。(3)通過圖位克隆和等位性測驗(yàn)的方法,我們克隆了 IPE1基因,可能編碼一種葡萄糖-甲醇-膽堿氧化還原酶。RT-PCR,qRT-PCR和原位雜交實(shí)驗(yàn)顯示IPE1基因特異地在幼嫩的雄穗中表達(dá),且在四分體和單核小孢子前期的花藥絨氈層中表達(dá)量高。(4)生化分析顯示突變體ipe1花藥脂肪酸顯著減少;與之相對應(yīng)地是,RNA-Seq數(shù)據(jù)分析說明部分參與脂肪酸合成和延長、蠟質(zhì)合成和類黃酮代謝的基因的表達(dá)被改變。暗示著IPE1通過參與脂類物質(zhì)的合成,從而調(diào)控花藥角質(zhì)層和花粉外壁的形成。(5)表型分析和表達(dá)分析證明:IPE1,MS26,MS45協(xié)同調(diào)控玉米花藥發(fā)育。IPE1和MS26參與花藥角質(zhì)層和花粉外壁的形成,MS45參與花藥角質(zhì)層的成熟和花粉外壁的增厚。(6)進(jìn)化樹和擬南芥T-DNA插入系分析顯示IPE1同源基因在雄性生殖器官發(fā)育中具有部分保守的功能。綜上所述,IPE1是ω-羥基脂肪酸氧化途徑中關(guān)鍵調(diào)節(jié)因子,與MS26和MS45協(xié)同調(diào)控玉米花藥角質(zhì)層和花粉外壁發(fā)育。我們的研究結(jié)果不僅為玉米雄性不育化制種提供了重要的基因資源;同時也豐富了玉米脂類物質(zhì)合成的遺傳網(wǎng)絡(luò)。
[Abstract]:Zea mays is an important food and cash crop. Male sterile lines have important application value in maize hybrid seed production. Anther cuticle and pollen outer wall are two protective structures of pollen formation. In Arabidopsis thaliana (Arabidopsis) and Oryza sativa (Oryza sativa), some genes controlling the development of anther cuticle and pollen outer wall have been cloned. However, only a few genes have been cloned in maize, and their molecular mechanisms are unclear. A male sterile line, -ipe1, was obtained by screening the library of Mutator mutants in laboratory. The IPE1 gene was cloned by map cloning and molecular biology, and the molecular mechanism of its regulation of maize fertility was preliminarily clarified. The main results were as follows: (1) phenotypic analysis showed that ipe1 had only abnormal anther development, while the other organs had no significant changes, compared with wild type, ipe1 anther could not emerge from glume, become smaller and wilt. Iodine-potassium iodide staining showed that ipe1 anthers had no mature pollen grain formation. (2) Cytology analysis showed that the formation of keratinocytes and urnytes in ipe1 anthers was blocked, and meiosis of ipe1 anthers was normal. The abnormal development of columnar layer resulted in random accumulation of sporopollen and irregular outer wall of pollen grains, which led to the gradual degradation of microspore from the middle stage of mononuclear stage, and the tapetum was degraded earlier than wild type by means of map cloning and allelic test. We cloned the IPE1 gene, which may encode a glucose-methanol-cholinergic redox enzyme. RT-PCRnqRT-PCR and in situ hybridization experiments showed that IPE1 gene was specifically expressed in young male ears. Biochemical analysis in anther tapetum of tetrad and premononuclear microspore showed that ipe1 anther fatty acids were significantly decreased, whereas RNA-Seq data showed that they were partly involved in fatty acid synthesis and prolongation. The expression of waxy synthesis and flavonoid metabolism genes was altered. Suggesting that IPE1 is involved in the synthesis of lipids, Thus the phenotypic analysis and expression analysis on the regulation of anther keratinocytes and pollen outer wall showed that 1: IPE1 and MS26 MS45 were involved in the development of maize anther. IPE1 and MS26 were involved in the formation of anther cuticle and pollen outer wall. MS45 was involved in the maturation of anther cuticle. Phylogenetic tree and Arabidopsis T-DNA insertion line analysis showed that IPE1 homologous gene had a partially conserved function in male reproductive organ development. In conclusion, IPE1 is a key regulatory factor in 蠅 -hydroxyl fatty acid oxidation pathway, and coordinated with MS26 and MS45 to regulate the development of horniness layer and pollen outer wall of maize anther. Our results not only provide important genetic resources for maize male sterility seed production, but also enrich the genetic network of lipid synthesis in maize.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：S513

【引證文獻(xiàn)】

相關(guān)期刊論文 前1條

1 宗緒巖;李麗;刁沖;王飛;;玉米威士忌生產(chǎn)工藝研究[J];四川理工學(xué)院學(xué)報(自然科學(xué)版);2017年05期

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本文編號：1832680